This invention claims priority of the German patent applications 100 30 013.8 and 101 15 577.8 which are incorporated by reference herein.
The invention relates to a scanning microscope with multiband illumination. In particular, the invention relates to a scanning microscope with an objective, through which a sample can be illuminated and detected, the objective being arranged in both an illumination beam path and a detection beam path, and an illumination pinhole and a detection pinhole being respectively arranged in the illumination beam path and in the detection beam path.
The invention furthermore relates to an optical component for a scanning microscope with multiband illumination.
European Patent EP-0 495 930 discloses a confocal microscope system for multicolour fluorescence. A dichroic mirror, which transmits a plurality of wavelength ranges, is provided in the detection beam path and observation beam path of the microscope. The dichroic mirror disclosed here is, however, configured only for specific wavelength ranges. Should the need arise that it is desirable to carry out studies with another combination of wavelength ranges, then it will be necessary to replace the dichroic mirror with one that is suitable for this combination of wavelength ranges. This considerably restricts the flexible usability of the instrument.
German laid-open patent specification 198 29 954 discloses a beam splitter in a laser scanning microscope. The beam splitter in the beam path of the laser scanning microscope is used to separate the excitation beam path and emission beam path. The beam splitter preferably consists of coloured glasses which can be changed. So that the coloured glasses can be changed, they are arranged on a splitter revolver and are rotated into the beam part according to use. This device has the disadvantage, however, that readjustment is often necessary in the case of changeable optical elements in the beam path of a laser scanning microscope.
It is an object of the present invention to provide a scanning microscope which makes it possible to study a sample with various wavelengths and/or wavelength ranges, while obviating the need to readjust wavelength-specific elements in the beam path.
The object is achieved by a scanning microscope comprising: an objective through which a sample can be illuminated and detected, the objective being arranged in both an illumination beam path and a detection beam path, an illumination pinhole being arranged in the illumination beam path, a detection pinhole being arranged in the detection beam path, an optical component arranged in the illumination beam path, which generates spectrally broadened illumination light, and an essentially polarization-independent and wavelength-independent beam splitter, which is arranged in a fixed position in the illumination beam path and the detection beam path.
It is another object of the invention to provide an optical component which contains, firmly mounted, all the elements that are sensitive with respect to adjustment, in order hence to permit straightforward replacement.
The object is achieved by an optical component for use in an illumination and detection beam path of a scanning microscope comprising: a polarization-independent and wavelength-independent beam splitter, an illumination pinhole, a detection pinhole and a reference detector.
It is advantageous to combine an optical component with a casing, and to fit it as a module in the beam path of a microscope, preferably a scanning microscope. The optical component comprises a beam splitter, an illumination pinhole, a detection pinhole and a reference detector. The optical component is arranged in the illumination beam path and the detection beam path of the scanning microscope in such a way that the illumination light is incident with respect to the normal of the beam splitter at an angle which is such that polarization effects are minimal.
It is furthermore advantageous if at least one adjustment means is respectively assigned to the beam splitter, the illumination pinhole and the detection pinhole.
Added to this, the optical component can be used in a scanning microscope. The scanning microscope comprises a light source, which consists of at least one laser and a microstructured material, into which the light from the laser can be injected. It has been shown that it is particularly advantageous if a reference detector is arranged in a reference beam defined by the light passing through the beam splitter. The reference detector derives from the light of the reference beam an electronic value that can be sent to control electronics. According to the electronic value, the intensity of the light source is controlled in such a way that there is always a constant light power at a sample.
Another advantage is that the optical component is enclosed by a casing. This very substantially prevents incorrect positioning of and/or damage to the individual elements in the casing. The casing comprises a first opening for entry of the illumination light, a second opening for exit of the illumination light and for entry of the detection light, and a third opening for exit of the detection light. The illumination pinhole is assigned to the first opening and the detection pinhole is assigned to the third opening. Furthermore, the beam splitter is arranged in a fixed position with respect to the illumination beam path and the detection beam path.
Other advantageous configurations of the invention can be found in the dependent claims.